1. Field of the Invention
The present invention relates to the field of nanotechnology. More particularly, the present invention provides nanoscale assembly systems for systemic delivery of therapeutic bioactive lipid compounds and/or hydrophobic chemotherapeutic agents and/or nucleotide/gene agents to individuals in need of such therapy.
2. Description of Related Art
Nanotechnology has been intricately linked with the life sciences (generally referred to as nanobiotechnology) since its inception by Richard Feynman in his 1959 speech, “There's Plenty of Room at the Bottom,” in which he made reference to the complexity and smallness of the living cell and challenged the scientific community to “make a thing very small which does what we want” (Feynman, R. P., 1959). Although commercial nanobiotechnology is still in its infancy, the rate of nanoscale assembly system development has been increasing exponentially in the last ten years, due to the unique advantages that these systems offer for drug delivery and therapeutics. Examples of some nanoscale assembly systems include liposomes, polymeric structures such as dendrimers and hydrogels, and metal or semiconductor nanoparticles referred to as quantum dots.
Many effective reagents are available for introducing transcriptionally active DNA, and even functional peptides and proteins into viable cells. However, approaches to deliver bioactive lipids into living cells are not generally available. The delivery of bioactive sphingolipids and phospholipid metabolites, analogues, mimetics or derivatives and their intercalation into cells is impeded by their physical-chemical properties that render these lipids hydrophobic and cell impermeable.
Ceramide, a sphingolipid that acts as a lipid-derived second messenger that modulates the induction of cell differentiation, cell cycle arrest and/or apoptosis, is an example of a bioactive lipid whose exogenous administration has been problematic. Intracellular ceramide accumulation results from multiple stimuli, such as growth factor deprivation, cytokines, chemotherapy and other cytotoxic agents, ionizing radiation, heat shock, and various environmental factors. These stimuli have been observed to initiate ceramide-mediated signaling cascades, including the inhibition of Akt pro-survival pathways and the stimulation of caspase activity, which ultimately leads to DNA fragmentation and cell death. Thus, based on ceramide's potent regulation of cell growth, differentiation, and death, and the fact that it is a natural molecule that targets discrete kinases and signaling pathways linked to proliferation and/or survival, ceramide has been identified as a therapeutic agent in cancer and cardiovascular disease.
The clinical utility of local delivery of a cell-permeable ceramide analogue, C6, from drug-eluting platforms previously has been demonstrated by Charles et al. (Circ. Res. 2000 Aug. 18:87(4):282-8). Specifically, ceramide-coated balloon catheters were shown to induce cell cycle arrest in stretch-injured vascular smooth muscle cells. Although the delivery of C6-ceramide from coated and distended balloons allow for direct delivery to the vasculature, there are several obstacles to the delivery of ceramide for systemic applications, such as cancer chemotherapy or targeting diffuse atherosclerotic lesions and vulnerable plaque. In particular, three significant barriers to systemic ceramide delivery exist, despite the use of short chain, more cell permeable derivatives.
First, short-chain, cell-permeable ceramide analogues such as C2, C6, and C8-ceramide are still lipids, and thus extremely hydrophobic by nature, precipitating as fine lipid micelle suspensions when added, in DMSO or ethanol vehicle, to cell media. Second, although short-chain ceramide analogues are more cell-permeable than long-chain physiological ceramide (C18-C24-ceramide), their sphingoid backbone limits their intercalation into plasma membranes. Finally, the existence of circulating and intracellular ceramidases promote the conversion of bioactive ceramides into less pro-apoptotic metabolites.
Organic solvent systems have been investigated in order to augment the delivery of ceramide to cells. It has been proposed that a dodecane/ethanol solvent system, which is insoluble in culture media, precipitates out with the ceramide and forms very small droplets, or micelles, that fuse with the plasma membrane. The use of such precipitating solvents may be limited by the variability in particle size and access to cellular membranes.
Protein adjuvants, such as bovine serum albumin, may also assist in vitro ceramide delivery via non-specific lipid/protein interactions, but would not permit the efficient delivery of sufficient quantities of C6-ceramide to systemic targets.
Thus, in order to realize the therapeutic benefits of bioactive lipids or gene therapy agents, there exists a need for improved systemic delivery systems of such hydrophobic or charged chemotherapeutic compounds into living cells of animals or humans in need of such therapy.